New Pyrroline Isolated from Antarctic Krill-Derived Actinomycetes Nocardiopsis sp. LX-1 Combining with Molecular Networking

Antarctic krill (Euphausia superba) of the Euphausiidae family comprise one of the largest biomasses in the world and play a key role in the Antarctic marine ecosystem. However, the study of E. superba-derived microbes and their secondary metabolites has been limited. Chemical investigation of the secondary metabolites of the actinomycetes Nocardiopsis sp. LX-1 (in the family of Nocardiopsaceae), isolated from E. superba, combined with molecular networking, led to the identification of 16 compounds a–p (purple nodes in the molecular network) and the isolation of one new pyrroline, nocarpyrroline A (1), along with 11 known compounds 2–12. The structure of the new compound 1 was elucidated by extensive spectroscopic investigation. Compound 2 exhibited broad-spectrum antibacterial activities against A. hydrophila, D. chrysanthemi, C. terrigena, X. citri pv. malvacearum and antifungal activity against C. albicans in a conventional broth dilution assay. The positive control was ciprofloxacin with the MIC values of <0.024 µM, 0.39 µM, 0.39 µM, 0.39 µM, and 0.20 µM, respectively. Compound 1 and compounds 7, 10, and 11 displayed antifungal activities against F. fujikuroi and D. citri, respectively, in modified agar diffusion test. Prochloraz was used as positive control and showed the inhibition zone radius of 17 mm and 15 mm against F. fujikuroi and D. citri, respectively. All the annotated compounds a–p by molecular networking were first discovered from the genus Nocardiopsis. Nocarpyrroline A (1) features an unprecedented 4,5-dihydro-pyrrole-2-carbonitrile substructure, and it is the first pyrroline isolated from the genus Nocardiopsis. This study further demonstrated the guiding significance of molecular networking in the research of microbial secondary metabolites.


Introduction
The extreme environments of Antarctica, including severe cold, an arid climate, and solar radiation, have created a unique ecological system. Creatures in the Antarctic ecosystem, in particular microorganisms, usually have to produce some structurally specific active substances to adapt to the harsh conditions [1]. Antarctic krill (Euphausia superba), a small crustacean in the family of Euphausiidae in the Antarctic Ocean, with one of the largest biomasses (approximately 379 million metric tons) in the world, plays a key role in the Antarctic marine ecosystem [2]. Antarctic krill is critical in the food chain for seals, whales, and penguins, making it the foundation of the Southern Ocean ecosystem and an important marine living resource [3,4]. The research on Antarctic krill has mainly focused on its fisheries [5], nutritive value [6], and distribution [7]. To the best of our knowledge, only three studies have focused on Antarctic krill-derived microbes and their secondary The molecular networking was used to preliminarily investigate the number and the structural types of the secondary metabolites of the actinomycetes Nocardiopsis sp. LX-1 and predicted the possibility of finding new compounds to determine the research value of the actinomycetes. After analysis by molecular networking, 16 compounds a-p (purple nodes) (Figures 2 and 3, Table 1) were characterized from the crude extracts of Nocardiopsis sp. LX-1 by comparing their MS/MS spectra with those in the GNPS library. The plastic products were avoided during the extraction and purification, and the plasticizers i and j might be contaminated by the organic extracted solvent, which were not the LC-MS grade/HPLC grade solvents that might contain contaminants. All the compounds a-p, identified by molecular networking, were preliminarily detected from the crude extracts of Nocardiopsis sp. LX-1. The big cluster families of the LX-1 molecular network with heavy molecular weight are speculated to be fatty acid compounds which could be lost in separated process. A series of antimicrobial isoflavonoids and flavonoids (a-f) were identified by molecular networking, which should be the target natural products to be isolated from LX-1. None of the recognized compounds a-p had been isolated from the genus Nocardiopsis until now, indicating the ability of molecular networking to analyze microbial secondary metabolites and guide the directional separation. The molecular networking was used to preliminarily investigate the number and the structural types of the secondary metabolites of the actinomycetes Nocardiopsis sp. LX-1 and predicted the possibility of finding new compounds to determine the research value of the actinomycetes. After analysis by molecular networking, 16 compounds a-p (purple nodes) (Figures 2 and 3, Table 1) were characterized from the crude extracts of Nocardiopsis sp. LX-1 by comparing their MS/MS spectra with those in the GNPS library. The plastic products were avoided during the extraction and purification, and the plasticizers i and j might be contaminated by the organic extracted solvent, which were not the LC-MS grade/HPLC grade solvents that might contain contaminants. All the compounds a-p, identified by molecular networking, were preliminarily detected from the crude extracts of Nocardiopsis sp. LX-1. The big cluster families of the LX-1 molecular network with heavy molecular weight are speculated to be fatty acid compounds which could be lost in separated process. A series of antimicrobial isoflavonoids and flavonoids (a-f) were identified by molecular networking, which should be the target natural products to be isolated from LX-1. None of the recognized compounds a-p had been isolated from the genus Nocardiopsis until now, indicating the ability of molecular networking to analyze microbial secondary metabolites and guide the directional separation.       Chemical investigation of the broth fermentative crude extracts of Nocardiopsis sp. LX-1 was carried out to find the molecular networking analyzed compounds, and led to the isolation of one new compound, nocarpyrroline A (1), along with 11 known compounds 2-12 ( Figure 4). However, only one target flavonoid derivative, daidzein (2, same as the molecular networking identified isoflavonoid e), in the molecular network was isolated, which might be due to the insufficient fermentation of Nocardiopsis sp. LX-1 and its low yield of the isoflavonoid and flavonoid compounds. This is the first time isoflavonoid has been isolated from the genus Nocardiopsis. Compounds 1, 2, and 4-8 were appeared in the molecular network of Nocardiopsis sp. LX-1 as individual nodes ( Figure 5). The self-loops of compounds 1 and 2 in the network might because their MS/MS spectra are not informative due to the low amount of the compounds; therefore no peak matches with other clustered nodes. Compounds 4-8 were appeared as individual nodes might be due to the fact that cyclic dipeptides are harder to be ionized in the positive ESI MS/MS experiment than in the negative ESI MS/MS experiment. Compound 9 was not detected in ESI + MS/MS spectra of Nocardiopsis sp. LX-1 because it cannot ionize in positive MS/MS measurement. Compounds 8 and 10-12 were not found in the molecular network might be due to their low molecular weight, which is hard to be detect. Mar. Drugs 2023, 21, x FOR PEER REVIEW 6 of 17   (Table 2 and Figure S1), combined with the six 13 Figure S2), indicated that there was a phenyl group in 1. The HMBC correlations from H-8 to C-9/C-10/C-14 proved the phenyl group was linked at C-8 ( Figure 6). The 1 H NMR, 13 Figure 6). One unsaturated quaternary carbon at δC 164.2 showed there was a carbonyl group in 1. The HMBC correlation between H-8 and C-6, and the COSY cross peak of H-7/H-8, revealed the carbonyl group was in the location of C-6 ( Figure 6). The high field    (Table 2 and Figure S1), combined with the six 13 Figure S2), indicated that there was a phenyl group in 1. The HMBC correlations from H-8 to C-9/C-10/C-14 proved the phenyl group was linked at C-8 ( Figure 6). The 1 H NMR, 13 Figure 6). One unsaturated quaternary carbon at δC 164.2 showed there was a carbonyl group in 1. The HMBC correlation between H-8 and C-6, and the COSY cross peak of H-7/H-8, revealed the carbonyl group was in the location of C-6 ( Figure 6). The high field  (Table 2 and Figure S1), combined with the six 13 Figure S2), indicated that there was a phenyl group in 1. The HMBC correlations from H-8 to C-9/C-10/C-14 proved the phenyl group was linked at C-8 ( Figure 6). The 1 H NMR, 13 C NMR, HSQC and HR-ESI-MS spectra (Figures S1-S3 and Figure S7) of 1 displayed one unsaturated quaternary carbon at δ C 135.2, one unsaturated methine at δ H 5.73 (1H, d, 2.7 Hz), δ C 119.6, one oxygenated methine at δ H 4.71 (1H, ddd, 8.2, 3.5, 2.7 Hz), δ C 70.2, and one methylene at δ H 3.72 (1H, dd, 13.6, 8.2 Hz), 3.62 (1H, dd, 13.6, 3.5 Hz), and δ C 54.5, demonstrated a pyrroline ring with a hydroxyl substituent group in 1. The COSY cross-peaks of H-3/H-4 and H-4/H-5, and the HMBC correlations from H-3 to C-2 and H-3 to C-5 further proved the existence of a pyrroline ring in 1 ( Figure 6). One unsaturated quaternary carbon at δ C 164.2 showed there was a carbonyl group in 1. The HMBC correlation between H-8 and C-6, and the COSY cross peak of H-7/H-8, revealed the carbonyl group was in the location of C-6 ( Figure 6). The high field shift of the C-6 carbonyl group indicated the amide linkage between C-6 and N-1  Table 2). The IR absorption band at 2253 cm -1 ( Figure S9), combined with the molecular formula of C 14 H 14 O 3 N 2 and the nine degrees of unsaturation suggested that there was one cyano group in 1. The high field shift of C-2 and low field shift of C-3 indicated that the cyano group was linked at C-2 (Table 2). Thus, the plane structure of 1 was established unambiguously as shown in Figure 4. The NOESY spectrum was measured to determine the relative configuration of 1; however, the NOESY cross-peaks were not clear enough to identify the relative configuration of 1. The absolute configuration of 1 was attempted to determine by the modified Mosher's method. Unfortunately, it was failed due to the limited quantity of 1. shift of the C-6 carbonyl group indicated the amide linkage between C-6 and N-1 ( Table  2). The IR absorption band at 2253 cm -1 ( Figure S9), combined with the molecular formula of C14H14O3N2 and the nine degrees of unsaturation suggested that there was one cyano group in 1. The high field shift of C-2 and low field shift of C-3 indicated that the cyano group was linked at C-2 (Table 2). Thus, the plane structure of 1 was established unambiguously as shown in Figure 4. The NOESY spectrum was measured to determine the relative configuration of 1; however, the NOESY cross-peaks were not clear enough to identify the relative configuration of 1. The absolute configuration of 1 was attempted to determine by the modified Mosher's method. Unfortunately, it was failed due to the limited quantity of 1.  The new compound nocarpyrroline A (1) was proposed to be biosynthesized by the condensation reaction of hydroxyl-cinnamate and 4-hydroxy-4,5-dihydro-1H-pyrrole-2carbonitrile ( Figure 7). Hydroxyl-cinnamate was suggested to be obtained from the hydroxylation of cinnamate. Phenyllactic acid (PLA) was biosynthesized from phosphoenolpyruvate and erythrose-4-phosphate through the enzymes of 3-deoxy-7-phosphoheptulonate synthase (DAHPS), 3-dehydroquinic acid synthase (DHQS), 3-dehydroquinic acid dehydratase (DHQD), shikimic acid 5-dehydrogenase (SDH), shikimic acid kinase (SK), 3-enolpyruvylshikimic acid 5-phosphate synthase (ESPS), chorismic acid synthase (CS), chorismic acid mutase (CM), prephenic acid aminotransferase (PAT), arogenic acid dehydratase (ADT), aminotransferase (ATF), dehydrogenase (DHG) in sequence [29,30]. Then, 4-hydroxy-4,5-dihydro-1H-pyrrole-2-carbonitrile was deduced to be achieved through amination, dehydration, hydroxylation, and reduction of L-Pro. L-Pro was biosynthesized from L-glutamate-γ-semialdehyde catalyzed by Δ 1 -pyrroline-5-carboxylate reductase. L-glutamate-γ-semialdehyde could be acquired by Δ 1 -pyrroline-5-carboxylate The new compound nocarpyrroline A (1) was proposed to be biosynthesized by the condensation reaction of hydroxyl-cinnamate and 4-hydroxy-4,5-dihydro-1H-pyrrole-2carbonitrile ( Figure 7). Hydroxyl-cinnamate was suggested to be obtained from the hydroxylation of cinnamate. Phenyllactic acid (PLA) was biosynthesized from phosphoenolpyruvate and erythrose-4-phosphate through the enzymes of 3-deoxy-7-phosphoheptulonate synthase (DAHPS), 3-dehydroquinic acid synthase (DHQS), 3-dehydroquinic acid dehydratase (DHQD), shikimic acid 5-dehydrogenase (SDH), shikimic acid kinase (SK), 3-enolpyruvylshikimic acid 5-phosphate synthase (ESPS), chorismic acid synthase (CS), chorismic acid mutase (CM), prephenic acid aminotransferase (PAT), arogenic acid dehydratase (ADT), aminotransferase (ATF), dehydrogenase (DHG) in sequence [29,30]. Then, 4-hydroxy-4,5-dihydro-1H-pyrrole-2-carbonitrile was deduced to be achieved through amination, dehydration, hydroxylation, and reduction of L-Pro. L-Pro was biosynthesized from L-glutamate-γ-semialdehyde catalyzed by ∆ 1 -pyrroline-5-carboxylate reductase. L-glutamate-γ-semialdehyde could be acquired by ∆ 1 -pyrroline-5-carboxylate synthase from L-Glu or received through catalyzing L-ornithine by ornithine-δ-aminotransferase (Figure 7) [31].
Compound 4 was acquired as an amorphous white powder. Its molecular formula was decided by HR-ESI-MS spectra (Figures S20 and S21)  , which was similar with those of 3. It was deduced that 4 was a DKP compound similar to 3. This deduction was further confirmed by the fact that the NMR data of 4 displayed two amido-carbonyl signals at δ C 169.8 and 165.5, and two amino-methine signals at δ H 4.07, dd (9.9, 6.4), δ C 58.5 and δ H 3.77, dd (5.7, 3.9), δ C 62.9 (Tables S2 and S3). Compound 4 was finally proved to be cyclo(4-methyl-D-Pro-L-Nva) (4) Its 1 H NMR, 13 C NMR, and specific OR data (Tables S2-S4) are almost identical to examples in the literature [36].
Compound 6 was obtained as a colorless oil. Its molecular formula was determined as C 14  . The only obvious difference between the NMR data of 6 and 5 was the methylene at C-4 in 6 was substituted by hydroxy-methine in 5. We carefully compared the NMR and specific OR data of 6 (Tables S2-S4) with those in the literature [38] and came to the determination that 6 is cyclo(L-Pro-L-Phe).
Compound 7 was acquired as a colorless oil. The HR-ESI-MS spectra ( Figures S32 and S33 Na, 249.1210) which was similar to those of 4. We deduced that 8 was a DKP compound similar to 4. This deduction was further confirmed by the NMR data of 8, which displayed two amido-carbonyl signals at δ C 168.8 and 172.9, and two amino-methine signals at δ H 4.50, ddd (11.2, 6.5, 1.6), δ C 58.5 and δ H 4.15, ddd (6.6, 4.4, 1.8), δ C 54.9 (Tables S2 and S3). Compound 8 was finally determined to be cyclo(4-hydroxyl-L-Pro-L-Leu) because it has almost the same 1 H NMR, 13 C NMR, and specific OR data (Tables S2-S4) as examples in the literature [40].
Compound 9 was obtained as an amorphous white powder. Its molecular formula was determined to be C 8 H 6 O 2 N 2 by HR-ESI-MS ( Figure S40) (Table S5), which were the NMR characteristic of quinazolinedione and almost same as those in the literature [41], so 9 was identified as 2,4(1H, 3H)quinazolinedione.
Compound 10 was acquired in the form of colorless needles. The HR-ESI-MS ( Figures S43 and S44) (Table S5) determined 10 to be uracil [42].

General Experimental Procedures
The UHPLC-MS/MS spectrum was obtained on a high-resolution Q-TOF mass spectrometry Bruker impactHD (Bruker, Switzerland, Germany), combined with Ulti-mate3000 UHPLC (Thermo Fisher Scientific, Waltham, MA, USA). A Thermo Scientific LTQ Orbitrap XL spectrometer (Thermo Fisher Scientific, Bremen, Germany) was used to measure HR-ESI-MS. Implen Gmbh NanoPhotometer N50 Touch (Implen, Munich, Germany) was used to record the UV spectrum. Nicoiet 380 (Thermo Fisher, Waltham, MA, USA) was used to measure the IR spectrum. Optical rotations were measured on a JASCO P-1020 digital polarimeter (JASCO, Tokyo, Japan). NMR spectra were measured on JEOL JNM-ECZ400S (JEOL, Tokyo, Japan). The Waters 1525 system was used for HPLC purification. Silica gel (200-300 mesh) was employed for chromatographic separation. Thinlayer chromatography was recorded on precoated silica gel GF254 plates.

Actinomycic Materials
The actinomycetes Nocardiopsis sp. LX-1 was isolated from the Antarctic krill Euphausia superba provided by Qingdao Dongfeng Ocean Fishing Co. LTD in 2019. The strain was deposited in the State Key Laboratory of Microbial Technology, Institute of Microbial Technology, Shandong University, Qingdao, China.
The identification of the actinomycetes LX-1 was determined by the analysis of the 16S rDNA gene sequence in NCBI (Nucleotide BLAST: Search nucleotide databases using a nucleotide query (nih.gov)). The 16S rDNA gene sequence of LX-1 was obtained through the polymerase chain reaction (PCR) method. The fresh actinomycetes LX-1 (about 1.00 mg) was dispersed in a 50-μL lysis buffer (Takara, Cat# 9164) and then was saved in a metal bath (Yooning, Hangzhou, China) at 100 °C for 30 min to extract its genomic DNA as the template DNA in PCR. The PCR was conducted in a final volume of 50 μL, which was composed of the template DNA (3 μL), primers 27F (1 μL) and 1492R (1 μL), PrimeSTAR ® Max DNA Polymerase (25 μL, Takara, Cat# R045A), and ultrapure water (20 μL), under the following procedures: (1) initial denaturation at 98 °C for 5 min, (2)

General Experimental Procedures
The UHPLC-MS/MS spectrum was obtained on a high-resolution Q-TOF mass spectrometry Bruker impactHD (Bruker, Switzerland, Germany), combined with Ultimate3000 UHPLC (Thermo Fisher Scientific, Waltham, MA, USA). A Thermo Scientific LTQ Orbitrap XL spectrometer (Thermo Fisher Scientific, Bremen, Germany) was used to measure HR-ESI-MS. Implen Gmbh NanoPhotometer N50 Touch (Implen, Munich, Germany) was used to record the UV spectrum. Nicoiet 380 (Thermo Fisher, Waltham, MA, USA) was used to measure the IR spectrum. Optical rotations were measured on a JASCO P-1020 digital polarimeter (JASCO, Tokyo, Japan). NMR spectra were measured on JEOL JNM-ECZ400S (JEOL, Tokyo, Japan). The Waters 1525 system was used for HPLC purification. Silica gel (200-300 mesh) was employed for chromatographic separation. Thin-layer chromatography was recorded on precoated silica gel GF254 plates.

Actinomycic Materials
The actinomycetes Nocardiopsis sp. LX-1 was isolated from the Antarctic krill Euphausia superba provided by Qingdao Dongfeng Ocean Fishing Co. LTD in 2019. The strain was deposited in the State Key Laboratory of Microbial Technology, Institute of Microbial Technology, Shandong University, Qingdao, China.
The identification of the actinomycetes LX-1 was determined by the analysis of the 16S rDNA gene sequence in NCBI (Nucleotide BLAST: Search nucleotide databases using a nucleotide query (nih.gov)). The 16S rDNA gene sequence of LX-1 was obtained through the polymerase chain reaction (PCR) method. The fresh actinomycetes LX-1 (about 1.00 mg) was dispersed in a 50-µL lysis buffer (Takara, Cat# 9164) and then was saved in a metal bath (Yooning, Hangzhou, China) at 100 • C for 30 min to extract its genomic DNA as the template DNA in PCR. The PCR was conducted in a final volume of 50 µL, which was composed of the template DNA (3 µL), primers 27F (1 µL) and 1492R (1 µL), PrimeSTAR ® Max DNA Polymerase (25 µL, Takara, Cat# R045A), and ultrapure water (20 µL), under the following procedures: (1) initial denaturation at 98 • C for 5 min, (2) denaturation at 98 • C for 30 s, (3) annealing at 55 • C for 30 s, (4) extension at 72 • C for 1.5 min, and (5) final extension at 72 • C for 10 min. Steps (2)-(4) were repeated 35 times. The PCR product was submitted to BGI Genomics for sequencing (BGI, Qingdao, China). The sequence of LX-1 was searched in the NCBI nucleotide collection database through the BLAST program (Nucleotide BLAST: Search nucleotide databases using a nucleotide query (nih.gov)). The actinomycetes LX-1 was identified as Nocardiopsis sp. whose 969 base pair 16S sequence had 99.9% sequence identity to that of Nocardiopsis sp. E251 (MT533941). The sequence data have been submitted to GenBank with accession number OL687477.

MS/MS Parameters
MS/MS analyses were performed by high-resolution Q-TOF mass spectrometry by using a Bruker impactHD. The ESI source parameters were set as follows: capillary source voltage at 3500 V, positive-ion mode, drying-gas temperature at 200 • C, drying-gas flow rate at 4 L/min, and end plate offset voltage at 500 V. MS scans were recorded in full scan mode with a range of m/z 50−1500 (100 ms scan time), and the mass resolution was 40,000 at m/z 1222.

Molecular Network Analysis
The molecular network was formed by GNPS workflow (http://gnps.ucsd.edu, accessed on 29 November 2021) [18]. Bruker Daltonics was used to convert the UHPLC-MS/MS raw data file into .mzXML. The parameter settings of the molecular network were detailed in our previous research [22,23]. The results were visualized by using the software package Cytoscape 3.8.0 (Download from https://cytoscape.org/).

Extraction and Isolation
The actinomycetes Nocardiopsis sp. LX-1 was cultured in a NB liquid medium in 100 Erlenmeyer flasks (200 mL in each 500 mL flask) at 20 • C for 45 days. The mycelia were filtered from the broth by two layers of gauze. Then, the mycelia were first extracted by ethyl acetate (EA) three times (3 × 200 mL) and then with dichloromethane (DCM)/methanol (MA) (v/v, 1:1) three times (3 × 200 mL). The c broth was obtained through repeated extraction with EA (3 × 20 L). All of the fungal crude extracts were put together and evaporated to dryness under reduced pressure to provide a residue (2.2 g). The residue was subjected to silica gel column chromatography (CC) eluted with EA-petroleum ether (PE) (0-100%) and MA-EA (0-100%) to obtain four fractions (Fr. control, respectively. The antifungal activities against eight phytopathogenic fungi, Aspergillus niger, Alternaria alternata, Diaporthe citri, Fusarium fujikuroi, F. oxysporum, F. graminearum, F. proliferatum, and Colletotrichum sp., were assessed through the modified agar diffusion test method [46]. The isolated compounds to be tested were dissolved in acetone at a final concentration of 100 µM. The compound solving solution was transferred to a sterile filter disk (diameter 6 mm, each 20 µL), which was placed on the agar growth medium for the tested fungi. Prochloraz was used as the positive control with the test concentration of 100 µM. Acetone was used as negative control.

Conclusions
In summary, 16 compounds a-p were recognized from the metabolites of Antarctic krill (E. superba)-derived actinomycetes Nocardiopsis sp. LX-1 by the method of molecular networking. One new pyrroline, nocarpyrroline A (1), along with 11 known compounds 2-12, were isolated from the actinomycetes Nocardiopsis sp. LX-1 according to the molecular networking analysis. Among them, compound 2 was the same as the molecular networking investigated isoflavonoid e, and this is the first time isoflavonoid from the genus Nocardiopsis has been isolated. New compound 1 showed antibacterial activity against A. hydrophila, and antifungal activity against F. fujikuroi. Compound 2 exhibited broad-spectrum antibacterial activities against A. hydrophila, D. chrysanthemi, C. terrigena, and X. citri pv. malvacearum, and antifungal activity against C. albicans. Compounds 7 and 10 displayed antibacterial activities against A. hydrophila, and 7, 10, and 11 revealed antifungal activities against D. citri. None of the annotated compounds a-p by the method of molecular networking had been isolated from the genus Nocardiopsis. Nocarpyrroline A (1), features an unprecedented 4,5-dihydro-pyrrole-2-carbonitrile substructure, and it is the first pyrroline discovered from the genus Nocardiopsis. This study further demonstrated the potential of Antarctic microbes to produce new bioactive natural products and proved the significance of molecular networking in the research of microbial secondary metabolites.
The extremely cold, arid, and fierce solar radiational environments of Antarctica, have created a unique ecological system containing abundant microbial resources with the ability to produce structurally specific active substances. Antarctic krill, as the foundation of the Antarctic marine ecosystem, contain rich symbiotic or parasitical microorganisms that produce special secondary metabolites. Few research papers have studied Antarctic krill-derived microorganisms and their secondary metabolites until now. More attention should be paid to chemical investigation and bioactive evaluation of the natural products isolated from Antarctic krill-derived microorganisms, which could find new bioactive compounds to provide the structural basis for new drug development.

Data Availability Statement:
The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number can be found below: https://www. ncbi.nlm.nih.gov/nuccore/OL687477.1/, accessed on 7 December 2021. The molecular network of the secondary metabolic profile of the actinomycetes Nocardiopsis sp. LX-1 can be found in https://gnps.ucsd.edu/ProteoSAFe/status.jsp?task=37dd96194c924f6d9daeef62672ba930, assessed on 29 November 2021.